Project Description/Abstract The targeting of chromatin modifiers is an increasingly attractive novel strategy to therapeutically inhibit transcription factors. MYC, one of the most frequently amplified oncogenes in cancer, is one such transcription factor that has proved difficult to target directly. Pediatric MYC-amplified medulloblastoma is a devastating disease. 25% of all medulloblastoma harbor amplification of MYC-isoforms that result in activation of MYC pathways. These tumors exhibit resistance to standard therapies used to treat medulloblastoma and are characterized by rapid and rampant tumor progression. Strategies to inhibit MYC activation pathways are desperately needed in the clinic for children diagnosed with this disease. Recently, inhibition of the epigenetic readers, BET-bromodomain proteins, has been found to be effective in suppressing the growth of preclinical models of MYC driven tumors, including medulloblastoma. Indeed, early phase clinical trials involving BET-bromodomain inhibitors are planned for children with recurrent MYC-driven tumors including medulloblastoma. However, the precise mechanism of action of these inhibitors remains unclear. In addition, clinical experience with other novel small molecule inhibitors has revealed that cancers evolve to acquire resistance to targeted therapeutics. Characterizing resistance mechanisms allows for novel therapeutic strategies to be designed to overcome these mechanisms and increase clinical efficacy of targeted therapeutics. The goal of this proposal is to systematically characterize cancer cell evolution in response to BET-bromodomain inhibition. This project will shed insight on the mechanism of action of BET-bromodomain inhibitors and guide the development of combination therapies to optimize efficacy. Cancers have been shown to acquire genetic alterations to develop resistance to targeted therapeutics. However, the mechanisms by which cancers evolve to acquire resistance to inhibition of chromatin modifiers have not been determined. BET-bromodomain proteins regulate the transcription of genes key to determination of cell-identity and cell-state. BET-bromodomain inhibitors have been shown to alter cell-state and cell-identity. It is thus possible that changes in cell-state can influence sensitivity to BET-bromodomain inhibition and contribute to the development of resistance. This proposal will systematically characterize the resistance mechanisms to BET-bromodomain inhibition in MYC-amplified medulloblastoma. To achieve this, a number of novel methodologies will be applied to characterize the genomic and epigenomic alterations that contribute to the acquisition of resistance to BET-bromodomain inhibition. This proposal will determine whether the acquisition of resistance is predetermined, will identify specific alterations in genes that contribute to resistance and will explore how resistance to BET-bromodomain inhibition is influenced by This work will guide the development of therapeutic strategies that will increase the clinical efficacy of these drugs. The results will also provide insights about cancers evolution following inhibition of chromatin modifiers that are likely to have pleiotropic effects. The modulation of chromatin modifiers is likely to be relevant to multiple cancers across all lineages. The mechanism(s) through which resistance accrues has not yet been determined in any of these cancers. This project will lay the framework for the study of chromatin modifiers that can translate across these diseases. An Instructor in Pediatrics and a Pediatric Neuro-Oncologist, Dr. Pratiti Bandopadhayay is completing post- doctoral training in the laboratory of Dr. Rameen Beroukhim at the Dana-Farber Cancer Institute and the Broad Institute of MIT and Harvard. Building on her clinical expertise and strong background in cancer biology, Pratiti is gaining expertise in cancer genomics and epigenomics, with a specific focus on studying cancer evolution. Dr. Bandopadhayay will complete this project under the co-mentorship of Dr. Beroukhim and Dr. Myles Brown, both physician-scientists. Dr. Beroukhim, himself a neuro-oncologist, is an international expert of cancer genomics, in particular of copy-number variations in cancers. He has developed methodologies, including GISTIC to profile copy-number alterations in cancer. He also has considerable experience in studying cancer genomes from data generated by whole genome and whole exome sequencing, with an interest in studying cancer heterogeneity and evolution. His appointments at the Broad Institute and the Dana-Farber Cancer Institute ensure that Dr. Bandopadhayay will have access to a tremendous range of resources to develop the project and to apply novel genomic approaches. Dr. Brown is a leader of chromatin biology, and has extensive experience in mentoring physician scientists. Under their mentorship, with the guidance of a strong scientific advisory committee, Dr. Bandopadhayay has proposed an ambitious research and training program that will equip her with the highest research skills to ensure her success in the mentored and independent phases of the award. The rich research environment of Dana-Farber Cancer Institute and the Broad Institute, in combination with the mentorship team of Drs. Beroukhim and Brown will provide the perfect training environment for Dr. Bandopadhayay to further her scientific training to emerge as an independent physician- scientist in the field of Pediatric Neuro-Oncology.